Cannabis sativa is currently the most commonly-used, illicit drug in the United States. Cannabis use, particularly by adolescents, increases the risk of developing schizophrenia-like psychoses in later life. In addition, people with bipolar disorder have a 20-40% lifetime prevalence of cannabis use, compared to 6% in the general population. These and other epidemiological data demonstrate that cannabis use predisposes susceptible individuals to the development of psychiatric disorders. Glycogen synthase kinase 3 (GSK-3) is emerging as an important regulatory kinase in the limbic brain;over-activity of GSK-3 has been linked to both bipolar disorder and schizophrenia. GSK-3 is phosphorylated and inactivated by protein kinases, including Akt and is a down-stream component of several neurotransmitters involved in mood regulation and psychosis. Data presented in this proposal demonstrate that prolonged cannabinoid-1 receptor (CB1R) activation by the cannabis constituent, ?9-tetrahydrocannabinol (THC), significantly decreases GSK-3 phosphorylation in neurons. Since GSK-3 activity is reduced by phosphorylation, these data are consistent with enhanced GSK-3 activity following THC exposure. A second CB1R agonist, CP55940 shared these effects of THC. These preliminary data, together with data in the literature regarding D2 dopamine receptor signaling, were used to formulate the hypothesis that prolonged CB1R activation results in recruitment of ?-arrestin;?-arrestin functions as a scaffold protein, bringing Akt in proximity with the protein phosphatase, PP2A. Akt is dephosphorylated and inactivated, resulting in dysinhibition of GSK-3 activity. Since GSK-3 over-activity is associated with mood dysregulation and psychosis, these data lead to the hypothesis that THC-mediated increase in GSK-3 activity contributes to the relationship between cannabis use and psychiatric disorders. The objective of the current project is to test the specific hypothesis that CB1R-mediated activation of GSK-3 occurs through ?-arrestin-mediated inhibition of Akt;is brain region specific and contributes to the anxiogenic and stress-enhancing effects of THC and other cannabinoid agonists. The specific aims of this project are: (1) to determine the mechanism by which CB1R agonists alter the phosphorylation state of GSK-3 in primary neurons in culture;(2) the determine the effects of acute and chronic exposure of mice to CB1R agonists and antagonists on the phosphorylation and activities of Akt and GSK-3 in brain regions, particularly those of the limbic system: (3) to determine the role of ?-arrestin in the effects of CB1R agonists and antagonists in behavioral assays of anxiety and stress by comparing their effects in wild type and ?-arrestin-2 null mice. Successful completion of the studies outlined in this proposal will advance our long-term objective to determine the mechanisms by which cannabis exposure predisposes individuals to the development of psychiatric illness. PUBLIC HEALTH RELEVANCE: Cannabis sativa is an annual plant that has been used by humans for thousands of years as a medicinal and recreational drug. While most individuals who use cannabis are not adversely affected, some users develop serious psychiatric disorders, including schizophrenia and bipolar disorder. The studies in this proposal will test the hypothesis that the primary psychoactive chemical of cannabis, ?9-tetrahydrocannabinol (THC) produces an over-activation of the enzyme glycogen synthase kinase in the brain and that this mechanism contributes to negative effects of THC on mood and cognition and is responsible for the increased incidence of psychiatric disorders in cannabis users.